Project Summary/Abstract Dyregulation of the mitogen-activated protein kinase (MAPK) and phosphatidylinositide-3-kinase (PI3K) pathways has been implicated in breast cancer pathology. Activation of the MEK1/2-ERK1/2 MAPK and the PI3K/Akt cascades in breast cancer increases cell proliferation and sustains tumor survival. Further, interaction between these pathways has been implicated in chemoresistance, suggesting that dual inhibition of these pathways may be necessary to prevent breast cancer growth and metastasis. The newest member of the MAPK family of kinases, the MEK5-ERK5 pathway, has been shown to enhance cancer cell growth and increase patient mortality. More specifically, increased ERK5 activation has been associated with decreased disease-free survival in breast cancer patients and signaling crosstalk with the PI3K pathway has been demonstrated in other cancer models. Therapies that target the MEK1/2-ERK1/2 pathways are used clinically and inhibitors of the PI3K/Akt pathway are in Phase I and II clinical trials for breast cancer. While there are currently 3 small molecule inhibitors of MEK5-ERK5 signaling available only one has been tested in breast cancer models. Moreover, in some breast cancer cells, these inhibitors are not specific for the MEK5-ERK5 pathway and, in some cases, even increase ERK5 activation. Therefore, there is a need for novel small molecule inhibitors that are specific for the MEK5-ERK5 pathway that may be clinically useful alone or in combination with other kinase inhibitors. The goal of this proposal is to test small molecule inhibitors of MEK5 that we developed in in vitro and in vivo breast cancer models and to use these novel inhibitors and biochemical methods to examine crosstalk between the MEK5-ERK5 and PI3K/Akt pathways in breast carcinoma. Therefore, the aims of this proposal seek to test the hypothesis that novel small molecule inhibitors of MEK5-ERK5 decrease tumorigenesis, migration, and metastasis and enhance the anti-proliferative effects of PI3K/Akt inhibitors in breast cancer. Breast cancer cell lines with distinct hormonal profiles will be employed and a breast cancer-xenograph mouse model will be used to examine the effect of the MEK5 inhibitors on breast tumor progression and metastasis. The experiments proposed in this application will provide a greater mechanistic understanding of the MEK5-ERK5 pathway, potentially leading to the development of novel, targeted therapeutics for breast cancer patients. Further, a more comprehensive understanding of the MEK5-ERK5 pathway may have widespread implications for various cancer types, adding to the significance of the proposal. These aims will also provide a stimulating training opportunity for undergraduate and graduate students to actively participate in the research and discovery processes to improve our understanding of signal transduction in cancer biology.